Membrane Topology of the ATP Binding Cassette Transporter ABCR and Its Relationship to ABC1 and Related ABCA Transporters

Abstract

ABCR is a member of the ABCA subclass of ATP binding cassette transporters that is responsible for Stargardt macular disease and implicated in retinal transport across photoreceptor disc membranes. It consists of a single polypeptide chain arranged in two tandem halves, each having a multi-spanning membrane domain followed by a nucleotide binding domain. To delineate between several proposed membrane topological models, we have identified the exocytoplasmic (extracellular/lumen) N-linked glycosylation sites on ABCR. Using trypsin digestion, site-directed mutagenesis, concanavalin A binding, and endoglycosidase digestion, we show that ABCR contains eight glycosylation sites. Four sites reside in a 600-amino acid exocytoplasmic domain of the N-terminal half between the first transmembrane segment H1 and the first multi-spanning membrane domain, and four sites are in a 275-amino acid domain of the C half between transmembrane segment H7 and the second multi-spanning membrane domain. This leads to a model in which each half has a transmembrane segment followed by a large exocytoplasmic domain, a multi-spanning membrane domain, and a nucleotide binding domain. Other ABCA transporters, including ABC1 linked to Tangier disease, are proposed to have a similar membrane topology based on sequence similarity to ABCR. Studies also suggest that the N and C halves of ABCR are linked through disulfide bonds.ABCR, formerly known as the rim protein, is an ATP binding cassette (ABC) 1 transporter found in vertebrate retinal photoreceptor cells (1-3). It is localized along the rim region of photoreceptor rod outer segment disc membranes (1, 4, 5) and more recently has been found in human foveal and peripheral cone outer segments (6). Several studies have implicated ABCR in the retinoid cycle, possibly functioning as a retinal extruder or retinal-phosphatidylethanolamine flippase to facilitate the removal of all-trans-retinal from disc membranes following the photobleaching of rhodopsin (7-9).Mutations in the ABCA4 gene encoding ABCR have been linked to Stargardt disease, a relatively common juvenile macular dystrophy characterized by a decrease in visual acuity, progressive bilateral atrophy of the central retina, and the accumulation of yellow deposits within the retinal pigment epithelial cell layer (2, 10 -13). Mutations in ABCA4 have also been linked to related disease variants, including late-onset fundus flavimaculatus (14), cone-rod dystrophy (15), retinitis pigmentosa-like dystrophy (16, 17), and age-related macular degeneration (18), although the latter remains controversial (19).ABCR is a member of the ABCA subclass of ABC transporters (20). Like most other mammalian ABC transporters, members of this subclass consist of a single long polypeptide chain organized into two tandemly arranged halves. Each half contains a membrane-spanning domain (MSD) followed by a cytoplasmic nucleotide binding domain (NBD). The ABCA subclass is distinguished from other ABC transporter subclasses by the presence of a large domain between the...

ATP-binding cassette transporter ABCA4 and chemical isomerization protect photoreceptor cells from the toxic accumulation of excess 11- cis -retinal

“…ABCA4, also known as the Rim protein, is localized along the rim region of rod and cone outer segment disk membranes (25,26). This localization is dictated by the exocytoplasmic domains (ECD) of ABCA4 (27). These domains are too large to fit within the luminal space between the flattened disk membranes but can fit in the space outlined by the rim region (12,27,28).…”

“…This localization is dictated by the exocytoplasmic domains (ECD) of ABCA4 (27). These domains are too large to fit within the luminal space between the flattened disk membranes but can fit in the space outlined by the rim region (12,27,28). It remains to be determined if the transport and clearance of 11-cisand all-trans-retinal from photoreceptors is further facilitated by the rim localization of ABCA4.…”

Dominant Negative Mechanism Underlies Autosomal Dominant Stargardt-like Macular Dystrophy Linked to Mutations in ELOVL4

“…Endoglycosidase Digestion-The total membrane fractions of bovine retina and transfected HEK 293T cells were isolated as described previously (36). Membranes were treated with N-glycosidase F and endoglycosidase H (New England Biolabs, Beverly, MA) according to the manufacturer's instructions but with modifications.…”

An ATP-binding Cassette Transporter Is a Major Glycoprotein of Sea Urchin Sperm Membranes

“…This region is thought to be a regulatory region in ABCA1 (25), and several models have been proposed describing the topology of this region in subfamily A members (24,30). With ABCA1, data support a model for an intracellular location with the hydrophobic region (TMS 7) sticking into the membrane but not passing through it (24).…”